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5-Azacytidine (5-azaC) was originally identified as an anticancer drug (NSC102876) which can cause hypomethylation of tumor suppressor genes. To assess its effects on runt-related transcription factor 3 (RUNX3), expression levels and the promoter methylation status of the RUNX3 gene were assessed. We also investigated alteration of biologic behavior of esophageal carcinoma TE-1 cells. MTT assays showed 5-azaC inhibited the proliferation of TE-1 cells in a time and dose-dependent way. Although other genes could be demethylated after 5-azaC intervention, we focused on RUNX3 gene in this study. The expression level of RUNX3 mRNA increased significantly in TE-1 cells after treatment with 5-azaC at hypotoxic levels. RT-PCR showed 5-azaC at 50 μM had the highest RUNX3-induction activity. Methylation-specific PCR indicated that 5-azaC induced RUNX3 expression through demethylation. Migration and invasion of TE-1 cells were inhibited by 5-azaC, along with growth of Eca109 xenografts in nude mice. In conclusion, we demonstrate that the RUNX3 gene can be reactivated by the demethylation reagent 5-azaC, which inhibits the proliferation, migration and invasion of esophageal carcinoma TE-1 cells.

To determine the efficacy of postoperative adjuvant chemotherapy with paclitaxel plus cisplatin (Taxol+DDP, TP therapy) for stage IIA esophageal squamous cell carcinoma (ESCC) and to investigate the expression of RUNX3 in lymph node metastasis-negative esophageal cancer and its relationship with medical prognosis, a retrospective summary of clinical treatment of 143 cases of stage IIA esophageal squamous cell carcinoma patients was made. The patients were divided into two groups, a surgery alone control group (52 patients) and a chemotherapy group that received postoperative TP therapy (91 patients). The disease-free and 5 year survival rates were compared between the groups and a multivariate analysis of prognostic factors was performed. The same analysis was performed for cases classified as RUNX3 positive and negative, with post-operative specimens assessed by immunohistochemistry. Although the disease-free and 5 year survival rates in control and chemotherapy groups did not significantly differ and there was no significance in RUNX3 negative cases, postoperative adjuvant chemotherapy in the chemotherapy group was shown to improve disease-free and 5 year survival rate compared to the control group in RUNX3 positive cases. On Cox regression multivariate analysis, postoperative adjuvant chemotherapy (P

The aim of this study was to investigate the role of RUNX3 in esophageal squamous cell carcinoma (ESCC) cells biological behavior and the relationship between the expression of RUNX3 and MMP-9, TIMP-1, ICAM-1. RUNX3 levels in 90 esophageal squamous cell carcinoma specimens using immunohistochemical staining to examine the correlation between RUNX3 expression and clinical stage of ESCC. Furthermore, the role of RUNX3 in ESCC progression was evaluated in vitro by siRNA-mediated knockdown of RUNX3 or lentivirus-mediated over-expression of RUNX3 in ESCC cell lines. The expression and activities of MMP-9, TIMP-1, and ICAM-1 were analyzed. We found decreased expression of RUNX3 in ESCC tissue to be significantly related to T stage of tumor (p

Runt-related transcription factor 3 (RUNX3) is a tumor suppressor gene whose reduced expression may play an important role in the development and progression of esophageal squamous cell cancer (ESCC). The aim of this study was to investigate the clinical relevance of RUNX3 in ESCC patients and effects of overexpression on biological behaviour of Eca109 cells in vitro and in vivo. Immunohistochemistry was performed to detect the clinical relevance of RUNX3 and lymph node metastasis in 80 ESCC tissues and 40 non-cancerous tissues using the SP method. RT-PCR and Western blotting were applied to assess the RUNX3 level and verify the Eca109 cell line with stable overexpression. Localization of RUNX3 proteins was performed by cell immunofluorescence. CCK-8 and Scrape motility assays were used to determine proliferation and migration and the TUNEL assay to analyze cell apoptosis. Invasive potential was assessed in cell transwell invasion experiments. In nude mice, tumorigenesis in vivo was determined. Results showed decreased expression of RUNX3 in esophageal tissue to be significantly related to lymph node metastasis (LNM) ( P

The present study investigated the effects of 5-azacytidine (5-azaC) on the expression level of the human runt-related transcription factor 3 (RUNX3) gene and the biological behavior of esophageal carcinoma Eca109 cells. The effect of the demethylation reagent 5-azaC on the viability of Eca109 cells was detected by the MTT assay, which demonstrated that 5-azaC inhibited the viability of Eca109 cells in a time- and dose-dependent manner. Although demethylation of other genes may occur following treatment with 5-azaC, we focused on the RUNX3 gene. When treated with 5-azaC at hypoxic levels, the expression of RUNX3 increased and the methylation degree of the RUNX3 gene was decreased significantly in Eca109 cells. 5-azaC at 50 µM demonstrated the highest RUNX3-induction activity, inducing RUNX3 mRNA and protein expression, and decreasing the degree of methylation of the RUNX3 gene. Methylation specific PCR indicated that 5-azaC induced RUNX3 expression through demethylation. The abilities of migration and invasion of Eca109 cells were inhibited by 5-azaC. The growth of Eca109 cells treated with 5-azaC in vivo was detected by a tumorigenesis experiment. 5-azaC inhibited the growth of Eca109 xenografts in nude mice. Taken together, our findings demonstrated that the RUNX3 gene is hypermethylated in Eca109 cells and that 5-azaC induces the expression of the RUNX3 gene by demethylation, which inhibits the proliferation, migration and invasion of Eca109 cells.